In a groundbreaking achievement that promises to redefine our understanding of liver biology and disease modeling, researchers have successfully engineered a multi-zonal liver organoid from human pluripotent stem cells that closely mimics the liver’s complex spatial architecture. This innovation addresses a long-standing challenge in hepatic biology: replicating the liver’s zonal heterogeneity in vitro. For decades, scientists have recognized that hepatocytes, the liver’s primary functional cells, are organized into distinct subpopulations arrayed along the portal-central axis, each specialized to perform unique metabolic roles. However, replicating this intricate zonal pattern, fundamental for proper liver function and metabolic homeostasis, in a laboratory dish remained elusive—until now.
The liver’s architecture is zonally demarcated, with periportal regions near the portal vein, interzonal regions, and pericentral regions near the central vein, each harboring hepatocytes exhibiting distinct gene expression profiles and metabolic capabilities. This zonation governs critical pathways such as the urea cycle, glutathione synthesis, and glucose metabolism, and it profoundly influences how the liver responds to injury. Reza et al. took on the formidable task of recreating this spatial diversity by pioneering a self-assembling organoid system derived from human induced pluripotent stem cells (hiPSCs). Through carefully designed preconditioning strategies involving well-known bioactive molecules—namely ascorbate and bilirubin—they were able to coax hepatic progenitors into zone-specific phenotypes that spontaneously organized into a spatially ordered microtissue.
This method hinges on the concept that ascorbate and bilirubin, previously implicated in directing zonal hepatic fates in vivo, can be harnessed to steer the differentiation and spatial arrangement of hepatocyte-like cells in culture. By enriching distinct hepatic progenitor populations with these molecules prior to co-culture, the resulting three-dimensional organoids established clear gradients and spatial segregation resembling the portal-central axis of the liver. This represents the first in vitro human model that authentically recapitulates the multi-zonal hepatic architecture, opening new avenues for more physiologically relevant studies in liver biology, toxicity testing, and regenerative medicine.
To deeply characterize the cellular identities within these organoids, the researchers employed single-nucleus RNA sequencing (snRNA-seq), enabling high-resolution dissection of the gene expression landscapes across individual nuclei. This approach elucidated a hepatoblast differentiation trajectory that aligns with known periportal, interzonal, and pericentral hepatocyte populations in the human liver. The transcriptomic data demonstrated that not only do these cells exhibit hallmark gene expression signatures unique to their zonal identity, but they also organize coherently within the organoid, reinforcing the spatial and functional authenticity of the model.
Complementing the transcriptomic insights, epigenetic analyses revealed sophisticated regulatory mechanisms underpinning the establishment and maintenance of zonal identity. The study uncovered that ascorbate and bilirubin influence the binding of the histone acetyltransferase EP300 to distinct partners—TET1 or HIF1α respectively—modulating chromatin accessibility and gene expression patterns specific to each zonal phenotype. This discovery highlights a finely tuned molecular interplay where bioactive molecules dynamically shape the epigenetic landscape to direct hepatic zonation.
The functional validation of the zonally patterned hepatic organoids was equally compelling. Cells exhibited zone-specific metabolic activities—such as those related to the urea cycle and glutathione metabolism—corroborating their molecular zonation profiles. This level of functional fidelity is unprecedented in human organoid models and paves the way for more accurate modeling of liver metabolism and disease states that traditionally depend on zonal vulnerability.
Crucially, the translational potential of this technology was demonstrated through transplantation experiments in immunodeficient rats subjected to bile duct ligation, a model of liver injury and cholestasis. The multi-zonal human organoids improved survival outcomes in these animals by mitigating hyperammonaemia and hyperbilirubinaemia—two key pathological hallmarks of liver dysfunction. This proof-of-concept suggests that zone-specific organoids could form the basis of novel cell therapies or bioengineered grafts aimed at restoring liver function in patients with chronic liver diseases.
The implications of this work extend beyond regenerative medicine applications. The study provides an unprecedented platform to dissect the molecular mechanisms governing liver development, zonal specification, and disease pathogenesis. Researchers can now investigate how distinct hepatocyte subpopulations respond differently to toxins, infections, or metabolic stressors within a controlled human system, something that has been historically constrained by species differences and limitations of traditional cell culture systems.
Moreover, the establishment of a robust human multi-zonal organoid model heralds transformative possibilities in drug discovery. Pharmaceutical compounds often have zone-dependent hepatotoxicity profiles, which have been exceedingly difficult to predict in vitro. These organoids offer a sophisticated assay platform to screen for such liabilities early in the drug development pipeline, potentially reducing late-stage drug failures and enhancing safety assessments.
From a developmental biology perspective, the revealed interaction between small molecules like ascorbate and bilirubin with key epigenetic regulators uncovers new dimensions of hepatic zonation control. The dual role of EP300 partnering with either TET1 or HIF1α in the context of distinct metabolic milieus reflects a novel layer of metabolic-epigenetic crosstalk critical for cellular specialization along physiological gradients.
Looking ahead, the ability to engineer spatially complex human liver tissues opens exciting prospects for the study of hepatic diseases characterized by zonal dysfunction—such as non-alcoholic fatty liver disease, fibrosis, and viral hepatitis. Understanding how zone-specific injury and repair mechanisms unfold offers hope for developing targeted therapeutics that precisely modulate dysfunctional hepatocyte subsets.
In addition to advancing scientific understanding, this study exemplifies the power of integrating stem cell biology, single-cell genomics, and epigenetics to recapitulate human organ complexity in vitro. The self-assembling, multi-zonal liver organoid represents a major step toward fully functional bioartificial livers, with scalability and human relevance that surpass existing models.
By delivering an in vitro system that authentically mirrors the liver’s division of labor and spatial heterogeneity, Reza and colleagues have created a versatile platform with profound impact across multiple biomedical disciplines. This accomplishment not only deepens insight into liver biology but also spotlights the translational potential of pluripotent stem cell-derived organoids in tackling complex organ-level phenomena.
As organoid technology continues to evolve, these innovations underscore a future where human liver diseases can be modeled with unprecedented accuracy and where personalized medicine strategies may harness patient-specific, zonally organized liver tissues for therapeutic intervention. The confluence of stem cell engineering, molecular biology, and regenerative therapy embodied in this study is poised to accelerate the journey from bench to bedside in liver health and disease.
Subject of Research:
Human liver zonal architecture and hepatic progenitor differentiation using pluripotent stem cell-derived liver organoids
Article Title:
Multi-zonal liver organoids from human pluripotent stem cells
Article References:
Reza, H.A., Santangelo, C., Iwasawa, K. et al. Multi-zonal liver organoids from human pluripotent stem cells. Nature (2025). https://doi.org/10.1038/s41586-025-08850-1
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Tags: bioactive molecules in organoid developmenthepatic organoid engineeringhepatocyte zonal heterogeneityhuman pluripotent stem cellsliver architecture and functionliver biology and disease modelingliver injury response mechanismsmetabolic roles of hepatocytesmulti-zonal liver organoidsself-assembling organoid systemsstem cell-derived liver modelsurea cycle and liver metabolism